The invention relates generally to a mobile phase preparation device for use with liquid chromatography, in particular high pressure liquid chromatography (HPLC) or ultra-high pressure liquid chromatography (UHPLC). Basically, liquid chromatography (LC) is a physical method of separation in which the components to be separated are distributed between a mobile phase and a stationary phase, through or along which the mobile phase percolates in a definite direction.
The field of liquid chromatography started with the use of gravity pressure to move liquids through columns packed with a large particle solid phase support, such as with 20 to 50 micrometers average particle size, in order to separate molecules. In the 1970s, high pressure liquid chromatography was introduced to provide better resolution and faster analyses by using pumps to move liquids through columns packed with small particles (three to ten micrometers) at pressures up to 6,000 psi (˜4.1×107 Pascal). Over the past decade, ultra-high pressure liquid chromatography, pioneered by James W. Jorgenson and co-workers in the late 1990s (see, for instance, Anal. Chem. 1997, 69, 983-989), has become very popular, as it provides even higher resolution and throughput, but requires pumps that move liquids through columns packed with very small particles (less than two micrometers) at pressures up to 15,000 psi (˜1.0×108 Pascal) and more. In order to achieve the maximum benefits of UHPLC, instrumentation needed to be redesigned to withstand these higher pressures, but also to minimize system volumes which adversely impact resolution, throughput and reproducibility at UHPLC pressures.
Modern commercially available HPLC and UHPLC systems have a pressure transducer and a purge valve in the fluid path of each solvent delivering pump prior to combining the solvents in a manifold (see fluid schematic in FIG. 1). These components, especially the pressure transducer, can add a significant volume to the fluid path between the pump outlet and the manifold inlet, generally between 500 to 1,000 microliters. Although systems can use software to compensate for the compressibility of the different solvents used in each pump, for example by speeding up the pump to compress the solvent, this software compensation gets more difficult to do accurately as the volume of solvent to be compressed increases. At extremes of a gradient separation, characterized by continuous or stepwise change of the composition of the mobile phase during the elution process, such as 0 to 5 percent of one solvent and, correspondingly, 100 to 95 percent of another solvent in a binary system, the solvent from the higher flowing pump can actually flow into the compression region of the lower flowing pump to compress its solvent to the system pressure. This cross flow of solvent can result in inaccurate solvent gradients and/or result in longer re-equilibration times between runs to insure the proper solvent mixture is being proportioned into the mixer at the start of a gradient run.
Although many UHPLC system components have been designed with pre-combination solvent compression in mind, there has not been any previous development that combined multiple system components to properly address these issues.